Signal transduction via guanine nucleotide binding proteins (G proteins) is crucial in regulation of cardiovascular, neural, endocrine, and immune cell function as well as in cell growth and differentiation. It has been increasingly recognized that knowledge of the dynamic aspects of signal transduction is important to a full description of signaling mechanisms. This is most evident in the function of the novel family of C protein regulators (RGS proteins). They have been recognized only in the last few years but appear to be responsible for subsecond temporal responses of C protein regulated ion channels and for modulation of slower classical signal transduction pathways. While much in vitro biochemical information is available for RGS proteins, much less is known about their in vivo functions or their potential as targets of drug design. The three aims of this project will: 1) explore mechanisms and specificity of RGS proteins using biochemical and spectroscopic approaches, 2) test via a gene targeting approach the role of RGS proteins in the physiological actions of Gi family G proteins such as regulation of cardiac automaticity and opiate analgesia, tolerance, and dependence, and 3) examine the role and specificity of individual endogenous RGS proteins by use of ribozyme knock-down methods. The studies in this proposal should permit a better understanding of rapid G protein activation and deactivation mechanisms and will explore the potential of RGS proteins as a novel target of drug action.